TWiV 238Lost in transla1on

1

retroviruses

PoliovirusSindbis virus Influenza virus

VSV

reovirus

hepa77s B virus

2

5-untranslated region

3 >1,000 nt in length, typically 50 70 ntOFen contains RNA secondary structures; must be unwound to allow passage of ribosome

Length and secondary structure influence transla7on efficiency

3-untranslated region

Can regulate transla7on ini7a7on, transla7on efficiency, mRNA stability

poly(A) tail, necessary for efficient transla7on

3

Transla7onal machinery: Ribosomes

RNA=pink, yellowProtein=blue

4

Transla7onal machinery: tRNAs

5

Transla1onal machinery

Ribosomes tRNAs ini7a7on proteins (eIF) elonga7on proteins (eEF) termina7on proteins (eRF)

6

5-end dependent ini1a1on

7

8

9

Internal ini1a1on

10

IRES = internal ribosome entry site11

12

13

all eIFs

all eIFs except eIF4E

eIF2, eIF3

14

Viral IRESes Picornaviruses Flaviviruses (hepa77s

C virus) Pes7viruses (bovine

viral diarrhea virus, classical swine fever virus)

Retroviruses (SIV, MMLV, HTLV, FLV)

Insect picorna-like viruses (cricket paralysis virus, Plau7a stali intes7ne virus)

Cellular IRESes BiP c-Myc Antennapaedia Ornithine decarboxylase Fibroblast growth factor Vascular endothelial growth factor

protein kinase p58PITSLRE

15

AminoacylPep7dylExit

16

Methionine-independent ini1a1on

Can assemble 80S ribosomes without any eIFs or Met-tRNAi

RNA mimics tRNAi

17

18

Decoding the viral genome

Polyprotein (Picornaviridae, Flaviviridae, Togaviridae, Arenaviridae, Bunyaviridae, Retroviridae) Subgenomic mRNAs (Rhabdoviridae, Paramyxoviridae, Togaviridae) Segmented genome (Orthomyxoviridae, Reoviridae) Internal ini7a7on (IRES) (Picornaviridae, Flaviviridae) Leaky scanning (Retroviridae, Paramyxoviridae) Re-ini7a7on of transla7on (Orthomyxoviridae, Herpesviridae) Suppression of termina7on (Retroviridae, Togaviridae) Ribosomal frameshiFing (Retroviridae)

19

Polyprotein synthesis

20

Leaky scanning

21

Re-ini1a1on of transla1on

22

Suppression of termina1on

eRF1 and eRF3 recognize all 3 stop codons (UAA, UAG, UGA

But occasionally stop codons may be recognized by a charged tRNA

23

Suppression of termina1on

24

Ribosomal frameshiLing

25

26

Regula1on of transla1on in virus-infected cells

Ini7a7on Elonga7on Termina7on

27

28

eIF2 kinases

29

PKR

30

31

Adenovirus VA RNA I prevents ac1va1on of PKR

dsRNA

active

VA RNA I

eIF2 subunitphosphorylation

P P

inactive

PKR

inactive

no phosphorylation of eIF2 subunit

active protein synthesis

protein synthesis inhibited

32

Vaccinia virus encoded pseudosubstrates mimic eIF2

33

Targets of viral inhibitors of eIF2 phosphoryla1on

34

35